SODs are a family of enzymes that catalyze the destruction of superoxide ions. SODs are metalloproteins that are individually characterized by their respective metal ions--which can be iron, manganese, or copper and zinc. The ability of SODs to catalyze the destruction of superoxide ions renders them useful in a variety of therapeutic settings such as in reducing reperfusion injury and in treating inflammation. The amino acid sequence of human Cu/Zn SOD is described in Jabusch et al, Biochemistry (1980) 19:2310-2316. The cloning and sequencing of human Cu/Zn SOD and the production of human Cu/Zn SOD in bacteria and yeast are described in EPA 84111416.8 (published Apr. 24, 1985 under number 0138111). Human Cu/Zn SOD is normally a homodimer of two chains bound together by hydrophobic interaction. The homodimer has a molecular weight of approximately 32 kd.
The primary structure of human Mn SOD is described by Barra et al, J.B.C. (1984) 259:12595-12601. The cloning and sequencing of human Mn SOD and the production of human Mn SOD in yeast is described in copending U.S. application Ser. No. 918,534, filed Oct. 14, 1986.
In achieving its therapeutic potential SOD must (1) achieve access to the target tissue upon which it is to act (2) maintain its molecular integrity while in transit and at the site of action and (3) not be cleared too rapidly from circulation. Prior modifications of SOD have focused on only the latter two of these requirements. For example, commonly owned copending U.S. application Ser. No. 026,143, filed Mar. 16, 1987 describes SOD polymers composed of covalently coupled SOD dimers which are not cleared as rapidly from circulation as native SOD. Res Commun. in Chem. Path. & Pharmacol. (1980) 29:113-120 and Proc. Nat'l Acad. Sci. U.S.A. (1980) 77:1159-1163 describe conjugates of SOD and macromolecules such as polyethylene glycol that also have increased circulatory half-lives relative to wild-type SOD. J. Clin. Invest. (1984) 73:87-95 describes SOD encapsulated in liposomes to protect the SOD from proteolysis and extend its clearance time.
In contrast, the present invention is primarily directed to modifying SOD in a manner to enable it to more readily access the target tissue upon which it is to act and/or to extend its corporeal lifetime. This is achieved by modifying the amino acid sequence of SOD to include a heterologous peptide domain that binds to a moiety that localizes at the site where the superoxide dismutase therapy is desired or binds to the site itself. Such domains are sometimes referred to as "adhesive peptide signals" herein.
Prior workers have identified adhesive peptide signals of various proteins that bind to cells, bodies, or molecules that are real to or commonly localize at sites at which SOD therapy may be desirable. The signals for fibrinogen, fibronectin, and von Willebrand factor are described in Nature (1984) 309:30-33; Proc. Nat'l Acad. Sci. U.S.A. (1984) 81:4935-4939; Cell (1986) 44:517-518 and Cell (1987) 48:867-873. Signals for laminin are described in Science 1987) 238:1132, for extracellular superoxide dismutase (EC-SOD) in PNAS (1987) 84:6340, for platelet-derived growth factor (PDGF) in Nature (1986) 320:695, for tissue plasminogen activator (tPA) in Abstracts 1141, 1144, 1043, and 1587 in "Thrombosis and Haemostasis, Volume 58, Abstracts", Eleventh International Congress, Brussels, Belgium, July 1987, and chemotactic factor in PNAS (1987) 84:9233. Those for Antithrombin III are described in Abstracts 543-545, ibid, and in J. Biol. Chem. (1987) 262:8061, 11964, 17356. The adhesive peptide signals for anti-platelet GPIIb/IIIa receptor and anti-endothelial cell GPIIb/IIIa receptor antibodies are discussed in Abstracts 15, 715, 901 and 908 of "Thrombosis and Haemostasis, Volume 58, Abstracts", supra. Those for vitronectin appear in Abstracts 15-17 and 564, ibid., for urokinase in Abstract 1140, ibid., for thrombin inhibitor in Abstract 649, ibid. The signal for chemotactic inhibitor is disclosed in Science (1979) 203:461, and for chemotactic agonists in PNAS (1987) 84:7964. The signals for platelet factor-4 appear in Blood (1979) 53:604 and Biochem. J. (1980) 191:769 and those for Gamma IP10 in Nature (1985) 315:672.
Applicants are unaware of any prior efforts to incorporate any of these adhesive domains or signals into other polypeptides to impart binding activity thereto.